The ability of Mycobacterium tuberculosis to infect an extraordinary number of people, combined with the widespread emergence of multidrug resistance and opportunistic infections in AIDS, has lead to its placement on the NIAID list of biodefense and emerging infectious agents. M. tuberculosis persistence in human populations rests upon its ability to infect and survive in macrophages. Intracellular M. tuberculosis inhibits phagosomal maturation and resides in a pathogen-friendly phagosome escaping lysosomal bactericidal mechanisms and efficient antigen presentation. The long-term objectives of this project are to characterize mycobacterial products responsible for the M. tuberculosis phagosomal maturation block and identify the host cell membrane trafficking processes that are targeted by the mycobacterial factors. These phenomena have been linked to mycobacterial interference with membrane trafficking and organelle biogenesis processes controlled by host cell Rab GTPases. We hypothesize that M. tuberculosis interferes with specific Rab-interacting partners, including a Rab effector, phosphatidylinositol 3-kinase, that prepares phagosomes for tethering and fusion with other organelles. An integral part of this hypothesis is that M. tuberculosis lipids, which mimic mammalian phosphatidylinositols, affect organellar fusion and phagosomal maturation by interfering with phosphatidylinositol 3-kinase-dependent processes in the host cell. The specific aims are: 1. Identify M. tuberculosis lipid and protein products affecting phagosome maturation and characterize their mode of action. 2. Delineate the role of host cell membrane trafficking regulators including Rab5 effectors in M. tuberculosis phagosome maturation arrest. 3. Characterize novel, cell biology-based processes that can counteract M. tuberculosis phagosome maturation arrest. This three-prong approach will improve our understanding of a marquee pathogenic determinant of M. tuberculosis and provide a foundation for new interventions potentially combating both active disease and latent infection.